Technical Abstract:
Background: Salmonella enterica serovar Enteritidis (SE) is the only one of over 2000 serovars to frequently contaminate the contents of the egg and it is the world's leading cause of human salmonellosis. Some isolates produce high-molecular-mass LPS (HMM LPS) that alters avian reproductive tract biology in otherwise healthy hens. Cooperation between orally invasive biofilm forming strains with those that produce HMM LPS reliably produces high incidence egg contamination by contact infection. The objective of this research was to determine when subpopulations with reproductive tract tropism emerge during the infection pathway to the egg. Methods: Neutral sugar compositional data were derived by gas chromatography of derivatized lipopolysaccharide (LPS) obtained by hot water extraction from spleens of naturally infected mice, and from United Kingdom isolates. Cluster analysis combined with generation of bell curves was used for statistical analysis. In addition, hens were challenged with a mixture of strains to see which one would be recovered from eggs. Results: U. S. mouse isolates were most like U. S. avian intestinal samples, whereas United Kingdom isolates were most like avian reproductive tract and egg isolates. Non-reproductive tract organ isolates had significant loss of O-chain. Orally invasive biofilm forming isolates and strains HMM LPS produced different O-chain structures at the biofilm-permissive temperature of 25 but not at 37 C. Hens infected at a 91:9 positive/negative biofilm colony phenotype ratio yielded only the negative phenotype from eggs. Conclusions: These results indicate that the hen applies stringent selection pressure to subpopulations of SE and that only the strain with evidence of reproductive tract tropism makes it to the egg. The avian cecum was identified as the earliest in vivo environment that supported emergence of strains producing HMM LPS. These results suggest that diet and other factors that alter gut physiology should be investigated for their ability to specifically reduce egg contamination as compared to just achieving reduction in intestinal colonization.